b'Next-Generation Sequencing (NGS) | CE-IVD AssaysIGK Assay Next-Generation Sequencing (NGS) | CE-IVD AssaysLymphoTrack Dx IGK AssayAssay Description BackgroundThe LymphoTrack Dx IGK Assays for the IlluminaMiSeq, or ThermoThe LymphoTrack Dx IGK Assays represent a significant improvement Fisher Scientific Ion PGM and Ion S5 instruments are in vitroover existing fragment analysis clonality assays by providing two diagnostic products intended for next-generation sequencing (NGS)important and complementary uses:based determination of the frequency distribution of IGK gene rearrangements in patients suspected of having lymphoproliferative1.Detection of initial clonal populations.disease. These assays aid in the identification2. Identification of sequence information required to track clonalof lymphoproliferative disorders. rearrangements in subsequent samples.The human immunoglobulin kappa (IGK) gene locus on chromosome Summary and Explanation of the Test 2 (2p11.2) includes 76 V (variable) region genes spanning 7 subgroups and 5 J (joining) region gene segments upstream of the (C )region.In contrast to the IdentiClone fragment analysis assays for clonality that utilize two master mixes, these NGS assays contain a singleDuring lymphoid cell development, antigen receptor genes multiplex master mix to target conserved regions of the IGK gene locusundergo somatic gene rearrangements. Specifically, during B-cell described in lymphoid malignancies, thereby, reducing sample DNAdevelopment, genes encoding IGK molecules are assembled from requirements and simplifying the testing workflow. The LymphoTrackmultiple polymorphic gene segments that generate V-J combinations Dx IGK master mix primers are also designed with Illumina or Thermounique in both length and sequence.Fisher Scientific adapters and up to 24 different indices. This allowsIn addition, the kappa deleting element (K de ), approximately 24 kb amplicons generated from different indexed IGK master mixes to be pooled into a single library for loading onto one MiSeq flow cell,downstream of the J-C region can also rearrange with V gene Ion PGM or Ion S5 chips. segments and the isolated recombination signal sequence in theJ-C intron (J-C INTR).The associated LymphoTrack Dx Software is capable of sortingSince leukemias and lymphomas originate from the malignant complex NGS data by gene target, providing users the ability totransformation of individual lymphoid cells, all leukemias and reduce per sample testing costs by sequencing amplicons from anylymphomas generally share one or more cell-specific or clonal LymphoTrack Dx Assay (e.g. IGH, IGK, TRB, TRG) at the same time.antigen receptor gene rearrangements. Therefore, tests that detect In addition, the LymphoTrack Dx Software provides an easy andIGK clonal rearrangements can be useful in the study of B- cell streamlined method for visualization of data and guidelines providedmalignancies and complement IGH testing, as the in the instructions for use allow samples to be interpreted for evidenceIGK receptor is less susceptible to somatic mutations.or no evidence of clonality. Specimen Requirement Positive clonal and negative polyclonal DNA controls are included in kits.50 ng of high-quality genomic DNA. References1.EP Rock et al., J Exp Med 179 (1): 323-8 (1994).2. JE Miller et al., Molecular Diagnostic 4 (2): 102-117(1999).Simplified Representation of the IGK GeneDepicted are the variable region (V) or variable intragenic J-C intron (J-C INTR) genes involved Kdein IGK gene rearrangements in addition to the downstream consensus joining region genes (J) or Kdekappa deleting element (K de ).26'